Natural approach of using nisin and its nanoform as food bio-preservatives against methicillin resistant Staphylococcus aureus and E.coli O157:H7 in yoghurt.

BACKGROUND: Natural antimicrobial agents such as nisin were used to control the growth of foodborne pathogens in dairy products. The current study aimed to examine the inhibitory effect of pure nisin and nisin nanoparticles (nisin NPs) against methicillin resistant Staphylococcus aureus (MRSA) and E.coli O157:H7 during the manufacturing and storage of yoghurt. Nisin NPs were prepared using new, natural, and safe nano-precipitation method by acetic acid. The prepared NPs were characterized using zeta-sizer and transmission electron microscopy (TEM). In addition, the cytotoxicity of nisin NPs on vero cells was assessed using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The minimum inhibitory concentrations (MICs) of nisin and its nanoparticles were determined using agar well-diffusion method. Further, fresh buffalo's milk was inoculated with MRSA or E.coli O157:H7 (1 × 106 CFU/ml) with the addition of either nisin or nisin NPs, and then the inoculated milk was used for yoghurt making. The organoleptic properties, pH and bacterial load of the obtained yoghurt were evaluated during storage in comparison to control group. RESULTS: The obtained results showed a strong antibacterial activity of nisin NPs (0.125 mg/mL) against MRSA and E.coli O157:H7 in comparison with control and pure nisin groups. Notably, complete eradication of MRSA and E.coli O157:H7 was observed in yoghurt formulated with nisin NPs after 24 h and 5th day of storage, respectively. The shelf life of yoghurt inoculated with nisin nanoparticles was extended than those manufactured without addition of such nanoparticles. CONCLUSIONS: Overall, the present study indicated that the addition of nisin NPs during processing of yoghurt could be a useful tool for food preservation against MRSA and E.coli O157:H7 in dairy industry.

Occurrence, molecular characterization, and antimicrobial susceptibility of sorbitol non-fermenting Escherichia coli in lake water, fish and humans in central Oromia, Ethiopia.

Contaminated lake water and fish can be sources of bacterial pathogens of public health concern, including pathogenic E. coli. Within Ethiopia, specifically, Central Oromia, raw fish consumption is a common practice. Although there are few reports on occurrence of E. coli O157 in fish destined for human consumption and children under five years, information on the transmission pathways of E. coli O157 and other sorbitol non-fermenting (SN-F) E. coli from water-to-fish-to-human, and their virulence factors and antimicrobial resistant determinants along the fish supply chain is lacking. The study aimed to investigate the occurrence, molecular characteristics, and antimicrobial susceptibility of E. coli O157 and other SN-F E. coli strains in fish, lake water and humans in central Oromia, Ethiopia. A total of 750 samples (450 fish samples, 150 water samples, 150 human stool samples) were collected from five lakes and three health facilities. The samples were processed following the standard protocol recommended by European Food Safety Authority and Kirby-Bauer disc diffusion method for detection of the bacteria, and antimicrobial susceptibility tests, respectively. Molecular characterization of presumptive isolates was performed using Whole-Genome Sequencing (WGS) for serotyping, determination of virulence factors, antimicrobial resistance traits, and genetic linkage of the isolates. Overall, 3.9% (29/750) of the samples had SN-F E. coli; of which 6.7% (n = 10), 1.8% (n = 8) and 7.3% (n = 11) were retrieved from water, fish, and diarrheic human patients, respectively. The WGS confirmed that all the isolates were SN-F non-O157: H7 E. coli strains. We reported two new E. coli strains with unknown O-antigen from fish and human samples. All the strains have multiple virulence factors and one or more genes encoding for them. Genetic relatedness was observed among strains from the same sources (water, fish, and humans). Most isolates were resistant to ampicillin (100%), tetracycline (100%), cefotaxime (100%), ceftazidime (100%), meropenem (100%), nalidixic acid (93.1%) and sulfamethoxazole/trimethoprim (79.3%). Majority of the strains were resistant to chloramphenicol (58.6%) and ciprofloxacin (48.3%), while small fraction showed resistance to azithromycin (3.45%). Isolates had an overall MDR profile of 87.5%. Majority, (62.1%; n = 18) of the strains had acquired MDR traits. Genes encoding for mutational resistance and Extended-spectrum beta-lactamases (ESBL) were also detected. In conclusion, our study revealed the occurrence of virulent and MDR SN-F E. coli strains in water, fish, and humans. Although no genetic relatedness was observed among strains from various sources, the genomic clustering among strains from the same sources strongly suggests the potential risk of transmission along the supply chain at the human-fish-environment interface if strict hygienic fish production is not in place. Further robust genetic study of the new strains with unknown O-antigens, and the epidemiology of SN-F E. coli is required to elucidate the molecular profile and public health implications of the pathogens.

Selective bacteriophages reduce the emergence of resistant bacteria in bacteriophage-antibiotic combination therapy.

Escherichia coli O157:H7 is a globally important foodborne pathogen with implications for food safety. Antibiotic treatment for O157 may potentially contribute to the exacerbation of hemolytic uremic syndrome, and the increasing prevalence of antibiotic-resistant strains necessitates the development of new treatment strategies. In this study, the bactericidal effects and resistance development of antibiotic and bacteriophage monotherapy were compared with those of combination therapy against O157. Experiments involving continuous exposure of O157 to phages and antibiotics, along with genetic deletion studies, revealed that the deletion of glpT and uhpT significantly increased resistance to fosfomycin. Furthermore, we found that OmpC functions as a receptor for the PP01 phage, which infects O157, and FhuA functions as a receptor for the newly isolated SP15 phage, targeting O157. In the glpT and uhpT deletion mutants, additional deletion in ompC, the receptor for the PP01 phage, increased resistance to fosfomycin. These findings suggest that specific phages may contribute to antibiotic resistance by selecting the emergence of gene mutations responsible for both phage and antibiotic resistance. While combination therapy with phages and antibiotics holds promise for the treatment of bacterial infections, careful consideration of phage selection is necessary.IMPORTANCEThe combination treatment of fosfomycin and bacteriophages against Escherichia coli O157 demonstrated superior bactericidal efficacy compared to monotherapy, effectively suppressing the emergence of resistance. However, mutations selected by phage PP01 led to enhanced resistance not only to the phage but also to fosfomycin. These findings underscore the importance of exercising caution in selecting phages for combination therapy, as resistance selected by specific phages may increase the risk of developing antibiotic resistance.

Synergistic efficacy of ultrasound and ammonium persulfate in inactivating Escherichia coli O157:H7 in buffered peptone water and orange juice.

This study investigated the synergistic effects of ammonium persulfate (PS) and ultrasound (US) on the inactivation of Escherichia coli O157:H7 in buffered peptone water (BPW) and orange juice products. A comprehensive assessment of PS concentrations ranging from 1 to 300 mM, considering not only the statistical significance but also the reliability and stability of the experimental outcomes, showed that 150 mM was the optimal PS concentration for the inactivation of E. coli O157:H7. Additionally, US output intensities varying from 30 % to 60 % of the maximum US intensity were evaluated, and 50 % US amplitude was found to be the optimal US condition. A 50 % amplitude setting on the sonicator corresponds to half of its maximum displacement, approximately 60 µm, based on a maximum amplitude of 120 µm. The inactivation level of E. coli O157:H7 was significantly enhanced by the combined treatment of PS and US, compared to each treatment of PS and US alone. In the BPW, a 10-min treatment with the combination of PS and US resulted in a significant synergistic inactivation, achieving up to a log reduction of 3.86 log CFU/mL. Similarly, in orange juice products, a 5-min treatment with the combination of PS and US yielded a significant synergistic inactivation, with a reduction reaching 5.90 log CFU/mL. Although the treatment caused a significant color change in the sample, the visual differences between the treated and non-treated groups were not pronounced. Furthermore, the combined treatment in orange juice demonstrated significantly enhanced antimicrobial efficacy relative to BPW. Despite identical 5-min treatment periods, the application in orange juice resulted in a substantially higher log reduction of E. coli O157:H7, achieving 7.16 log CFU/mL at a reduced PS concentration of 30 mM, whereas the same treatment in BPW yielded only a 2.89 log CFU/mL reduction at a PS concentration of 150 mM, thereby highlighting its significantly superior antimicrobial performance in orange juice. The mechanism underlying microbial inactivation, induced by the combined treatment of PS and US, was identified as significant cell membrane damage. This damage is mediated by sulfate radicals, generated through the sono-activation of persulfate. In addition, the low pH of orange juice, measured at 3.7, is likely to have further deteriorated the E. coli O157:H7 cells compared to BPW (pH 7.2), by disrupting their cell membranes, proton gradients, and energy metabolism. These findings underscore the effectiveness of PS and US integration as a promising approach for non-thermal pasteurization in the food industry. Further research is needed to optimize treatment parameters and fully explore the practical application of this technique in large-scale food processing operations. Sensory evaluation and nutritional assessment are also necessary to address the limitations of PS.

Enhanced photodynamic inactivation against Escherichia coli O157:H7 provided by chitosan/curcumin coating and its application in food contact surfaces.

Sterilisation technologies are essential to eliminate foodborne pathogens from food contact surfaces. However, most of the current sterilisation methods involve high energy and chemical consumption. In this study, a photodynamic inactivation coating featuring excellent antibacterial activity was prepared by dispersing curcumin as a plant-based photosensitiser in a chitosan solution. The coating generated abundant reactive oxygen species (ROS) after light irradiation at 420 nm, which eradicated ≥99.999 % of Escherichia coli O157:H7. It was also found that ROS damaged the cell membrane, leading to the leakage of cell contents and cell shrinkage on the basis of chitosan. In addition, the production of ROS first excited the bacterial antioxidant defence system resulting in the increase of peroxidase (POD) and superoxide dismutase (SOD). ROS levels exceed its capacity, causing damage to the defence system and further oxidative decomposition of large molecules, such as DNA and proteins, eventually leading to the death of E. coli O157:H7. We also found the curcumin/chitosan coating could effectively remove E. coli O157:H7 biofilms by oxidative of extracellular polysaccharides and proteins. All the contributors made the chitosan/curcumin coating an efficient detergent comparable with HClO.

ß-Galactosidase-triggered in situ synthesis of yellow emitting silicon nanoparticle and its application in visual detection of E. coli O157:H7 and drug susceptibility test.

The sensitive detection of foodborne pathogenic and rapid antibiotic susceptibility testing (AST) is of great significance. This paper reports the enzyme-triggered in situ synthesis of yellow emitting silicon nanoparticles (SiNPs) and the detection of Escherichia coli (E. coli) O157:H7 in food samples and the rapid AST. The rapid counting of E. coli O157:H7 has been achieved through direct visual observation, equipment detection, and smartphone digitalization. A simple detection platform based on smartphone senses and cotton swabs has been established. Meanwhile, rapid AST based on enzyme-catalyzed SiNPs can intuitively obtain colorimetric samples. This paper established a system for bacterial enzyme-triggered in situ synthesis of SiNPs, with high responsiveness, luminescence ratio, and specificity. The detection limit for E. coli O157:H7 can reach 100 CFU/mL during 5 h, and the recovery efficiency ranges from 90.14% to 110.16%, which makes it a promising strategy for the rapid detection of E. coli O157:H7 and AST.

Ferrous sulfate combined with ultrasound emulsified cinnamaldehyde nanoemulsion to cause ferroptosis in Escherichia coli O157:H7.

The purpose of this study was to investigate ferroptosis in Escherichia coli O157:H7 caused by ferrous sulfate (FeSO4) and to examine the synergistic effectiveness of FeSO4 combined with ultrasound-emulsified cinnamaldehyde nanoemulsion (CALNO) on inactivation of E. coli O157:H7 in vitro and in vivo. The results showed that FeSO4 could cause ferroptosis in E. coli O157:H7 via generating reactive oxygen species (ROS) and exacerbating lipid peroxidation. In addition, the results indicated that FeSO4 combined with CALNO had synergistic bactericidal effect against E. coli O157:H7 and the combined treatment could lead considerable nucleic acids and protein to release by damaging the cell membrane of E. coli O157:H7. Besides, FeSO4 combined with CALNO had a strong antibiofilm ability to inhibit E. coli O157:H7 biofilm formation by reducing the expression of genes related on biofilm formation. Finally, FeSO4 combined with CALNO exhibited the significant antibacterial activity against E. coli O157:H7 in hami melon and cherry tomato.

Ferrous gluconate triggers ferroptosis in Escherichia coli: Implications of lipid peroxidation and DNA damage.

Microbial ferroptosis has been proved to combat drug-resistant pathogens, but whether this pattern can be applied to the prevention and control of Escherichia coli remains to be further explored. In this study, ferrous gluconate (FeGlu) showed remarkable efficacy in killing E. coli MG1655 with a mortality rate exceeding 99.9%, as well as enterotoxigenic E. coli H10407 (ETEC H10407) and enterohemorrhagic E. coli O157:H7 (EHEC O157:H7). Bacteria death was instigated by the infiltration of Fe2+, accompanied by a burst of intracellular reactive oxygen species (ROS) and lipid peroxidation. Notably, mitigating lipid peroxidation failed to alleviate death of E. coli. Further findings confirmed that FeGlu induced DNA damage, and ΔrecA mutant showed more sensitive, implicating that DNA damage was involved in the death of E. coli. The direct interaction of Fe2+ with DNA was demonstrated by fluorescent staining, gel electrophoresis, and circular dichroism (CD). Moreover, proteomic analysis unveiled 50 differentially expressed proteins (DEPs), including 18 significantly down-regulated proteins and 32 significantly up-regulated proteins. Among them, the down-regulation of SOS-responsive transcriptional suppressor LexA indicated DNA damage induced severely by FeGlu. Furthermore, FeGlu influenced pathways such as fatty acid metabolism (FadB, FadE), iron-sulfur cluster assembly (IscA, IscU, YadR), iron binding, and DNA-binding transcription, along with α-linolenic acid metabolism, fatty acid degradation, and pyruvate metabolism. These pathways were related to FeGlu stress, including lipid peroxidation and DNA damage. In summary, FeGlu facilitated ferroptosis in E. coli through mechanisms involving lipid peroxidation and DNA damage, which presents a new strategy for the development of innovative antimicrobial strategies targeting E. coli infections.

Bacteriological Survey of Fresh Minced Beef on Sale at Retail Outlets in Scotland in 2019: Three Foodborne Pathogens, Hygiene Process Indicators, and Phenotypic Antimicrobial Resistance.

ABSTRACT: The health and economic burden of foodborne illness is high, with approximately 2.4 million cases occurring annually in the United Kingdom. A survey to understand the baseline microbial quality and prevalence of food-related hazards of fresh beef mince on retail sale could inform risk assessment, management, and communication to ensure the safety of this commodity. In such a survey, a two-stage sampling design was used to reflect variations in population density and the market share of five categories of retail outlets in Scotland. From January to December 2019, 1,009 fresh minced beef samples were collected from 15 geographic areas. The microbial quality of each sample was assessed using aerobic colony count and Escherichia coli count. Samples were cultured for Campylobacter and Salmonella, and PCR was used to detect target genes (stx1 all variants, stx2 a to g, and rfbO157) for Shiga toxin-producing E. coli (STEC). The presence of viable E. coli O157 and STEC in samples with a positive PCR signal was confirmed via culture and isolation. Phenotypic antimicrobial sensitivity patterns of cultured pathogens and 100 E. coli isolates were determined, mostly via disk diffusion. The median aerobic colony count and E. coli counts were 6.4 × 105 (interquartile range, 6.9 × 104 to 9.6 × 106) and <10 CFU/g (interquartile range, <10 to 10) of minced beef, respectively. The prevalence was 0.1% (95% confidence interval [CI], 0 to 0.7%) for Campylobacter, 0.3% (95% CI, 0 to 1%) for Salmonella, 22% (95% CI, 20 to 25%) for PCR-positive STEC, and 4% (95% CI, 2 to 5%) for culture-positive STEC. The evidence for phenotypic antimicrobial resistance detected did not give cause for concern, mainly occurring in a few E. coli isolates as single nonsusceptibilities to first-line active substances. The low prevalence of pathogens and phenotypic antimicrobial resistance is encouraging, but ongoing consumer food safety education is necessary to mitigate the residual public health risk.

Logistic modeling to predict the minimum inhibitory concentration (MIC) of olive leaf extract (OLE) against Listeria monocytogenes.

Olive leaf extract (OLE) has been increasingly recognized as a natural and effective antimicrobial against a host of foodborne pathogens. This study attempts to predict the minimum inhibitory concentration (MIC) of OLE against Listeria monocytogenes F2365 by utilizing the asymptotic deceleration point (PDA) in a logistic model (LM), namely MIC-PDA. The experimental data obtained from the inhibitory rate (IR) versus OLE concentration against L. monocytogenes were sufficiently fitted (R2 = 0.88957). Five significant critical points were derived by taking the multi-order derivatives of the LM function: the inflection point (PI), the maximum acceleration point (PAM), the maximum deceleration point (PDM), the absolute acceleration point (PAA), and the asymptotic deceleration point (PDA). The PDA ([OLE] = 37.055 mg/mL) was employed to approximate the MIC-PDA. This MIC value was decreased by over 42% compared to the experimental MIC of 64.0 mg/mL, obtained using the conventional 2-fold dilution method (i.e., MIC-2fold). The accuracy of MIC-PDA was evaluated by an in vitro L. monocytogenes growth inhibition assay. Finally, the logistic modeling method was independently validated using our previously published inhibition data of OLE against the growths of Escherichia coli O157:H7 and Salmonella enteritidis. The MIC-PDA (for [OLE]) values were estimated to be 41.083 and 35.313 mg/mL, respectively, compared to the experimental value of 62.5 mg/mL. Taken together, MIC-PDA, as estimated from the logistic modeling, holds the potential to shorten the time and reduce cost when OLE is used as an antimicrobial in the food industry.

Control of Escherichia coli Serotype O157:H7 Motility and Biofilm Formation by Salicylate and Decanoate: MarA/SoxS/Rob and pchE Interactions.

Prophage-encoded Escherichia coli O157:H7 transcription factor (TF), PchE, inhibits biofilm formation and attachment to cultured epithelial cells by reducing curli fimbriae expression and increasing flagella expression. To identify pchE regulators that might be used in intervention strategies to reduce environmental persistence or host infections, we performed a computational search of O157:H7 strain PA20 pchE promoter sequences for binding sites used by known TFs. A common site shared by MarA/SoxS/Rob TFs was identified and the typical MarA/Rob inducers, salicylate and decanoate, were tested for biofilm and motility effects. Sodium salicylate, a proven biofilm inhibitor, but not sodium decanoate, strongly reduced O157:H7 biofilms by a pchE-independent mechanism. Both salicylate and decanoate enhanced O157:H7 motility dependent on pchE using media and incubation temperatures optimum for culturing human epithelial cells. However, induction of pchE by salicylate did not activate the SOS response. MarA/SoxS/Rob inducers provide new potential agents for controlling O157:H7 interactions with the host and its persistence in the environment. IMPORTANCE There is a need to develop E. coli serotype O157:H7 nonantibiotic interventions that do not precipitate the release and activation of virulence factor-encoded prophage and transferrable genetic elements. One method is to stimulate existing regulatory pathways that repress bacterial persistence and virulence genes. Here we show that certain inducers of MarA and Rob have that ability, working through both pchE-dependent and pschE-independent pathways.

New insights into the formation and recovery of sublethally injured Escherichia coli O157:H7 induced by lactic acid.

Escherichia coli O157:H7 can be injured by the action of lactic acid (LA) and injured cells can be recovered under suitable condition. In this study, RNA sequencing analysis revealed the overall genes change of sublethally injured (4 mM LA, 60 min; SI) and initial recovered (minA, 20 min; R) cells. Compared with untreated samples, 53 up-regulated and 98 down-regulated differentially expressed genes (DEGs; Padj < 0.05, change fold ≥2) were found in SI. Meanwhile, Genes related to carbohydrate transport and metabolic were up-regulated and the addition of carbohydrate increased cells resistance to LA. Genes involved in osmotic stress response and cell membrane integrity were down-regulated and E. coli O157:H7 cells were sensitive to osmotic stress during sublethal injury. Genes related to iron stress response and cation transport were changed and cation may affect sublethal injury formation by influencing production of ROS and cellular processes. In R, 1370 up-regulated and 1110 down-regulated DEGs were subdivided into various GO terms and membrane, biological adhesion, cell projection, oxidation-reduction process and catalytic activity, etc., showed significant enrichment (corrected P < 0.05). Particularly, genes related to fimbrial, flagellum and type III secretion system were up-regulated, which may improve infection ability and virulence property during recovery of injured cells. These findings provide novel insights into formation and recovery of sublethally injured E. coli O157:H7 induced by LA.

Synergistic inactivation of Escherichia coli O157:H7 and Staphylococcus aureus by gallic acid and thymol and its potential application on fresh-cut tomatoes.

Antibacterial activity against Escherichia coli O157:H7 and Staphylococcus aureus of five typical plant-derived compounds [gallic acid (G.A), citral (Cit), thymol (Thy), salicylic acid (S.A), lauric acid (L.A)] were investigated by determining the minimum inhibitory concentration (MIC) and the fractional inhibitory concentration index (FICI). The results showed that only a combination of Thy and G.A (TGA), with a concentration of 0.1 and 1.25 mg/mL, respectively, had a synergistic effect (FICI = 0.5) on both E. coli O157:H7 and S. aureus. The amount of Thy and G.A in mixture were four-fold lower than the MICs of the individuals shown to cause the equivalent antimicrobial activity in trypticase soy broth (TSB). The microbial reduction obtained in TSB with addition of TGA were significantly higher (P < 0.05) than the reduction shown for the broth supplemented with the separated phenolics. TGA caused the changes of morphology and membrane integrity of bacteria. Additionally, the application of TGA on fresh-cut tomatoes are investigated. Fresh-cut tomatoes inoculated with E. coli O157:H7and S. aureus were washed for 2min, 5min, 10min at 4 °C, 25 °C, 40 °C in 0.3% NaOCl, or water containing TGA at various concentrations. Overall, the reduction of TGA achieved against S. aureus is higher than E. coli O157:H7. Same concentrations of combined antimicrobials at a temperature of 40 °C further increased the degree of microbial inactivation, with an additional 0.89-1.51 log CFU/g reduction compared to that at 25 °C. Moreover, 1/2MICThy+1/2MICG.A at 25 °C for 10min or 40 °C for 5min were generally acceptable with sensorial scores higher than 7. Our results showed that TGA could work synergistically on the inactivation of the tested bacteria and may be used as an alternative disinfectant of fresh produce.

A Novel Nano-Antimicrobial Polymer Engineered with Chitosan Nanoparticles and Bioactive Peptides as Promising Food Biopreservative Effective against Foodborne Pathogen E. coli O157-Caused Epithelial Barrier Dysfunction and Inflammatory Responses.

For food quality and safety issues, the emergence of foodborne pathogenic bacteria has further accelerated the spread of antibiotic residues and drug resistance genes. To alleviate the harm caused by bacterial infections, it is necessary to seek novel antimicrobial agents as biopreservatives to prevent microbial spoilage. Nanoantimicrobials have been widely used in the direct treatment of bacterial infections. CNMs, formed by chitosan nanoparticles and peptides, are promising antibiotic alternatives for use as excellent new antibacterial drugs against pathogenic bacteria. Herein, the current study evaluated the function of CNMs in the protection of foodborne pathogen Escherichia coli (E. coli) O157 infection using an intestinal epithelial cell model. Antibacterial activity assays indicated that CNMs exerted excellent bactericidal activity against E. coli O157. Assessment of the cytotoxicity risks toward cells demonstrated that 0.0125-0.02% of CNMs did not cause toxicity, but 0.4% of CNMs caused cytotoxicity. Additionally, CNMs did not induced genotoxicity either. CNMs protected against E. coli O157-induced barrier dysfunction by increasing transepithelial electrical resistance, decreasing lactate dehydrogenase and promoting the protein expression of occludin. CNMs were further found to ameliorate inflammation via modulation of tumor factor α, toll-like receptor 4 and nuclear factor κB (NF-κB) expression via inhibition of mitogen-activated protein kinase and NF-κB activation and improved antioxidant activity. Taken together, CNMs could protect the host against E. coli O157-induced intestinal barrier damage and inflammation, showing that CNMs have great advantages and potential application as novel antimicrobial polymers in the food industry as food biopreservatives, bringing new hope for the treatment of bacterial infections.

Pleurotus eryngii polysaccharide nanofiber containing pomegranate peel polyphenol/chitosan nanoparticles for control of E. coli O157:H7.

Pomegranate peel polyphenols (PPP), which are natural, safe, and green antibacterial agents, were introduced and embedded in chitosan to form stable nanoparticles. The PPP@chitosan nanoparticles (PPP@CNPs) were further electrospun into nanofibers based on Pleurotus eryngii polysaccharide (PEP). The preferable distribution of particle size, polydispersity index, and zeta potential was realized through the addition of PPP at 3 mg/mL, which achieved the highest encapsulation rate of 23.71 ± 0.51%. The tensile strength and elongation at break of nanofibers reached 15.76 MPa and 0.69% with the addition of 1% PEP through electrospinning. The results of scanning electron microscopy (SEM) and atomic force microscopy (AFM) demonstrated that the addition of nanoparticles increased the diameter of PEP nanofibers from 148 nm to 163 nm, and the surface roughness of the fibers also increased. Meanwhile, the addition of nanoparticles improved the thermal stability of PEP nanofibers. PPP@CNPs/PEP nanofibers can inhibit the growth of E. coli O157:H7 on pork and cucumber surfaces during the five-days storage, and the inhibition rates were all above 95%. Besides, the nanofibers did not have any impact on the color and texture of foods.

Responses of Escherichia coli and Listeria monocytogenes to ozone treatment on non-host tomato: Efficacy of intervention and evidence of induced acclimation.

Because of the continuous rise of foodborne illnesses caused by the consumption of raw fruits and vegetables, effective post-harvest anti-microbial strategies are necessary. The aim of this study was to evaluate the anti-microbial efficacy of ozone (O3) against two common causes of fresh produce contamination, the Gram-negative Escherichia coli O157:H7 and Gram-positive Listeria monocytogenes, and to relate its effects to potential mechanisms of xenobiosis by transcriptional network modeling. The study on non-host tomato environment correlated the dose × time aspects of xenobiosis by examining the correlation between bacterial survival in terms of log-reduction and defense responses at the level of gene expression. In E. coli, low (1 µg O3/g of fruit) and moderate (2 µg O3/g of fruit) doses caused insignificant reduction in survival, while high dose (3 µg/g of fruit) caused significant reduction in survival in a time-dependent manner. In L. monocytogenes, moderate dose caused significant reduction even with short-duration exposure. Distinct responses to O3 xenobiosis between E. coli and L. monocytogenes are likely related to differences in membrane and cytoplasmic structure and components. Transcriptome profiling by RNA-Seq showed that primary defenses in E. coli were attenuated after exposure to a low dose, while the responses at moderate dose were characterized by massive upregulation of pathogenesis and stress-related genes, which implied the activation of defense responses. More genes were downregulated during the first hour at high dose, with a large number of such genes getting significantly upregulated after 2 hr and 3 hr. This trend suggests that prolonged exposure led to potential adaptation. In contrast, massive downregulation of genes was observed in L. monocytogenes regardless of dose and exposure duration, implying a mechanism of defense distinct from that of E. coli. The nature of bacterial responses revealed by this study should guide the selection of xenobiotic agents for eliminating bacterial contamination on fresh produce without overlooking the potential risks of adaptation.

In silico screening of ssDNA aptamer against Escherichia coli O157:H7: A machine learning and the Pseudo K-tuple nucleotide composition based approach.

This study was planned to in silico screening of ssDNA aptamer against Escherichia coli O157:H7 by combination of machine learning and the PseKNC approach. For this, firstly a total numbers of 47 validated ssDNA aptamers as well as 498 random DNA sequences were considered as positive and negative training data respectively. The sequences then converted to numerical vectors using PseKNC method through Pse-in-one 2.0 web server. After that, the numerical vectors were subjected to classification by the SVM, ANN and RF algorithms available in Orange 3.2.0 software. The performances of the tested models were evaluated using cross-validation, random sampling and ROC curve analyzes. The primary results demonstrated that the ANN and RF algorithms have appropriate performances for the data classification. To improve the performances of mentioned classifiers the positive training data was triplicated and re-training process was also performed. The results confirmed that data size improvement had significant effect on the accuracy of data classification especially about RF model. Subsequently, the RF algorithm with accuracy of 98% was selected for aptamer screening. The thermodynamics details of folding process as well as secondary structures of the screened aptamers were also considered as final evaluations. The results confirmed that the selected aptamers by the proposed method had appropriate structure properties and there is no thermodynamics limit for the aptamers folding.

Natural and synthetic plant compounds as anti-biofilm agents against Escherichia coli O157:H7 biofilm.

Escherichia coli is a common gram-negative bacterium found in the gut and intestinal tract of warm-blooded animals including humans. An evolved seropathotype E. coli O157:H7 (STEC) came into existence in 1982, since then it has been evolved as a stronger and more robust drug-resistant pathotype of E. coli. This drug resistance is due to horizontal gene transfer, natural gene evolution for survival, and most of the cases due to the ability of STEC to switch to the biofilm growth mode from planktonic lifestyle. During the growth in biofilm mode, Escherichia coli O157:H7 opts more robust ability to grow in adverse environments i.e., in presence of antibiotics and other antimicrobial chemicals. Due to the biofilm matrix, the microbial community acquires drug resistance. This makes the treatment of diseases caused by E. coli O157:H7 a complex challenge. To address the illnesses caused by this biofilm-forming pathogen, there are several possible strategies such as antibiotic therapies, synthetic antimicrobial chemicals, adjunct therapy of synergistic effect of multiple drugs, and more importantly plant originated compounds as a new anti-biofilm candidate. The present review summarizes various phytochemicals and their derivatives reported in the last decade mostly to eliminate the biofilm of STEC. The review will progressively reveal the antibiofilm mechanism of the phytochemicals against STEC and to be a potential candidate for the development of the future antibacterial drugs to STEC induced infections.

Chlorine inactivation of Escherichia coli O157:H7 in fresh produce wash process: Effectiveness and modeling.

Inactivation rate constant or inactivation coefficient (specific lethality) quantifies the rate at which a chemical sanitizer inactivates a microorganism. This study presents a modified disinfection kinetics model to evaluate the potential effect of organic content on the chlorine inactivation coefficient of Escherichia coli O157:H7 in fresh produce wash processes. Results show a significant decrease in the bactericidal efficacy of free chlorine (FC) in the presence of organic load compared to its absence. While the chlorine inactivation coefficient of Escherichia coli O157:H7 is 70.39 ± 3.19 L/mg/min in the absence of organic content, it drops by 73% for a chemical oxygen demand (COD) level of 600-800 mg/L. Results also indicate that the initial chlorine concentration and bacterial load have no effect on the chlorine inactivation coefficient. A second-order chemical reaction model for FC decay, which utilizes a proportion of COD as an indicator of organic content in fresh produce wash was employed, yielding an apparent reaction rate of (9.45 ± 0.22) × 10-4 /µM/min. This model was validated by predicting FC concentration in multi-run continuous wash cycles with periodic replenishment of chlorine.

Transition-Metal Dichalcogenide Artificial Antibodies with Multivalent Polymeric Recognition Phases for Rapid Detection and Inactivation of Pathogens.

Antibodies are recognition molecules that can bind to diverse targets ranging from pathogens to small analytes with high binding affinity and specificity, making them widely employed for sensing and therapy. However, antibodies have limitations of low stability, long production time, short shelf life, and high cost. Here, we report a facile approach for the design of luminescent artificial antibodies with nonbiological polymeric recognition phases for the sensitive detection, rapid identification, and effective inactivation of pathogenic bacteria. Transition-metal dichalcogenide (TMD) nanosheets with a neutral dextran phase at the interfaces selectively recognized S. aureus, whereas the nanosheets bearing a carboxymethylated dextran phase selectively recognized E. coli O157:H7 with high binding affinity. The bacterial binding sites recognized by the artificial antibodies were thoroughly identified by experiments and molecular dynamics simulations, revealing the significance of their multivalent interactions with the bacterial membrane components for selective recognition. The luminescent WS2 artificial antibodies could rapidly detect the bacteria at a single copy from human serum without any purification and amplification. Moreover, the MoSe2 artificial antibodies selectively killed the pathogenic bacteria in the wounds of infected mice under light irradiation, leading to effective wound healing. This work demonstrates the potential of TMD artificial antibodies as an alternative to antibodies for sensing and therapy.